Induction of Multiple Forms of Mouse Liver Cytochrome P-450. Evidence for Genetically Controlled De Novo Protein Synthesis in Response to Treatment with Beta-naphthoflavone or Phenobarbital

The administration of polycyclic aromatic compounds such as beta-naphthoflavone or 3-methylcholanthrene is known to cause the induction of many liver microsomal monoxygenase activities and the appearance of a distinct cytochrome called P-448 in genetically responsive, but not in nonresponsive, inbred mouse strains. However, the administration of 2,3,7,8-tetrachlorodibenzo-p-dioxin induces these activities and cytochrome P-448 formation to the same extent in both responsive and nonresponsive inbred strains. In contrast, phenobarbital or pregnenolone-16 alpha-carbonitrile induces in both responsive and nonresponsive strains a different profile of enzyme activities and the appearance of cytochrome P-450 (rather than cytochrome P-448). In the present studies, electrophoresis of liver microsomal proteins from inbred C57BL/6N and DBA/2N and recombinant inbred AKXL-38 and AKXL-38A mouse strains revealed the presence of four polypeptides whose relative staining intensity could be correlated with the induction state of the microsomes as determined by enzymatic and spectral methods. Of these four bands, Band 4 (55,000 daltons) was increased whenever spectral measurements revealed an increase in the cytochrome P-448 content due to administration of beta-naphthoflavone or 2,3,7,8-tetrachlorodibenzo-p-dioxin. Administration of pregnenolone-16alpha-carbonitrile caused an increase in Band 3 (54,000 daltons), whereas administration of phenobarbital caused an increase primarily in Band 2 (51,000 daltons) but also smaller increases in Band 1 (49,000 daltons) and Band 4. The changes observed for phenobarbital and pregnenolone-16alpha-carbonitrile were the same for both responsive and nonresponsive strains. The same electrophoretic technique was used to measure the incorporation of radioactive leucine into microsomal proteins. Microsomes were prepared from liver combined from responsive mice (C57BL/6N) treated with beta-naphthoflavone and L-[14C]leucine and nonresponsive mice (DBA/2N) treated with beta-naphthoflavone and L-[3H-4,5]leucine. A significant increase in the 14C/3H ratio was observed for Band 4, and decreases were seen for Bands 1 and 2. In similar experiments with other mice and phenobarbital as the inducing agent with L-[14C]leucine and the vehicle alone with L-[3H-4,5]leucine, the 14C/3H ratio was markedly increased for Band 2, and smaller increases were observed for Bands 1 and 4. These results and other data presented indicate that the increased formation of cytochrome P-448 and P-450 by beta-naphthoflavone and phenobarbital, respectively, is primarily the result of an increased rate of de novo protein synthesis rather than a decreased degradation rate or a conversion of pre-existing polypeptides.